The culture of microorganisms is in use in biological sciences, clinical sciences, and biotechnology. It makes use of microorganism culture devices.
Living cells are an example of microorganisms broadly used in biology. First, the most common culture method involves culture on the bottom of a Petri dish or T flask. With such devices, the presence of liquid and an uneven air-liquid/cell interface may impair the imaging of the cells when using e.g. upright microscopes. Cells can otherwise be observed using an inverted microscope as the bottom of the culture dish provides a well defined optical interface. Yet, at high magnification (>40×), the depth of focus becomes a concern. A thin coverslip is often placed over (upright microscope) or possibly below (inverted microscope) the cells. Placing the coverslip requires experimental skills in order not to e.g. squeeze cells, entrap a thick layer of liquid, or create leaks between the coverslip and the culture dish. Furthermore, samples with a coverslip cannot be reused. Therefore cells are difficult to observe with the most common culture methods.
Second, it might be advantageous to culture and study cells in microfluidics, as known in the art, because (i) it lowers costs for rare cells or cells difficult to culture, (ii) it enables better cell proliferation studies, (iii) cells have a better (smaller) ratio cells/volume of surrounding medium, and (iv) a faster exchange of nutrients and stimulating factors can be achieved. Some of these advantages are for instance described in A. Persidis, Nature Biotechnol., 1998, 16, 488-489.
In this respect, many microfluidic systems are made of polydimethylsiloxane (PDMS), due to its favorable mechanical properties, optical transparency, and bio-compatibility. Complex microfluidic cell culture devices have been for instance developed for diverse cell types such as Eukaryotic cells, lung cells, embryonic stem cells, and mammalian embryos. In particular, microfluidics are used for trapping and culturing cells: cells must be retained in specific areas that are suited for measurements; they may further need regular exchange of medium, controlled temperature and CO2. In addition, some critical cells (e.g. neurons) are difficult to culture in such systems because they need several days for attaching and developing a phenotype.
For the sake of exemplification thereof, let mention that US 2007/0090166 discloses a micro fluidic device which includes a substrate and membrane. The substrate includes a reservoir in fluid communication with a passage. A bio-compatible fluid may be added to the reservoir and passage. The reservoir is configured to receive and retain at least a portion of a cell mass. The membrane acts as a barrier to evaporation of the bio-compatible fluid from the passage. A cover may be added to cover the bio-compatible fluid to prevent evaporation of the bio-compatible fluid.
Next, US 2004/0265172 is directed to a microfluidic device for analyzing biological samples. The device is provided with a sample inlet section including an inlet port, a capillary passageway communicating with the inlet port and with an inlet chamber. The inlet chamber includes means for uniformly distributing the sample liquid across the inlet chamber and purging the air initially contained therein.
Somehow related, U.S. Pat. No. 7,138,270 discloses an assay device that comprises a base and glass plate lid. The base has an array of shallow microwells, each having a flat rim, all rims being co-planar. When the lid is placed on the base, a thin capillary gap is formed on each rim, acting as a liquid seal for a microwell chamber. The liquid is excess sample liquid and further excess is accommodated in overspill cavities between the microwells. Because of the liquid seal and shallow configuration the benefits of micro fluidic devices are achieved together with the handling convenience and use of conventional detection equipment of conventional microplate devices. As the lid is placed on the base, excess sample overspills into the surrounding overspill areas. A residual amount of sample fills a capillary gap between the rim and the lid. Therefore, the desired volume of the sample is completely surrounded by the base and the lid without any ambient air contact. The excess sample is on the rim forming the microchamber seal, and in the overspill cavities. Contact of the lid and the frames forms a second level of enclosure, also limiting access by ambient air and minimizing sample evaporation and contamination. The vents ensure uniform placement of the lid by allowing ambient air to escape as the overspill cavities receive excess sample during placement of the lid.
Also, U.S. Pat. No. 7,351,575 discloses a method for performing at least one bulk process step on a biological material comprising: a) chemically treating a base plate to enhance immobilization of kinase substrates thereon with a solution of mixed self-assembled monolayer (SAM) comprising about 0.1 to 20% maleimide-terminal groups in a background of tri(ethylene glycol) terminal groups; b) placing a first removable member on the base plate for establishing self-sealing contact of the first removable member on the base plate, the first removable member being adapted to repeatedly self-seal on the base plate, the first removable member further defining a plurality of first orifices therein, each having a plurality of first walls bounding respective ones of the plurality of first orifices, the first removable member further being configured such that, when placed in self-sealing contact with the base plate, the first removable member defines a plurality of first wells therewith corresponding to respective ones of the plurality of first orifices. The removable member is formed of a material capable of spontaneously forming a fluid-tight seal with surface when placed into contact therewith. A fluid-tight seal is achieved without the use of adhesives, ultrasound, heat or other means. The removable member is capable of being sealed to the surface, then removed (by means such as peeling and lifting, which may be performed manually or by machine) without damaging or leaving residue on the surface.
Beside the sole patent literature, a number of publications are devoted to the subject, amongst which a paper of Sung Jae Kim, et al., “Self-Sealed Vertical Polymeric Nanoporous-Junctions for High-Throughput Nanofluidic Applications”, ACS Publications. Here, the authors have developed an integration method of polymeric nanostructure in a poly(dimethylsiloxane) (PDMS)-based microfluidic channel, for nanofluidic applications. The polymer junction was created by infiltrating polymer solution between the gaps created by mechanical cutting, without any photolithography or etching processes. The PDMS can seal itself with the heterogeneous polymeric nanoporous material between the PDMS/PDMS gap due to its flexibility without any (covalent) bonding between PDMS and the polymer materials. Thus, it is possible to integrate the nanoporous junction into a PDMS microchip in a leak-free manner with repeatability.